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Applied and Environmental Microbiology, July 2002, p. 3352-3357, Vol. 68, No. 7
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.7.3352-3357.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Molecular Characterization and Expression of Pyruvate Formate-Lyase-Activating Enzyme in a Ruminal Bacterium, Streptococcus bovis

Narito Asanuma and Tsuneo Hino*

Department of Life Science, College of Agriculture, Meiji University, Higashimita, Tama-ku, Kawasaki 214-8571, Japan

Received 15 January 2002/ Accepted 10 April 2002

To clarify the significance of the activation of pyruvate formate-lyase (PFL) by PFL-activating enzyme (PFL-AE) in Streptococcus bovis, the molecular properties and gene expression of PFL-AE were investigated. S. bovis PFL-AE was deduced to consist of 261 amino acids with a molecular mass of 29.9 kDa and appeared to be a monomer protein. Similar to Escherichia coli PFL-AE, S. bovis PFL-AE required Fe2+ for activity. The gene encoding PFL-AE (act) was found to be polycistronic, and the PFL gene (pfl) was not included. However, the act mRNA level changed in parallel with the pfl mRNA level, responding to growth conditions, and the change was contrary to the change in the lactate dehydrogenase (LDH) mRNA level. PFL-AE synthesis appeared to change in parallel with PFL synthesis. Introduction of a recombinant plasmid containing S. bovis pfl and the pfl promoter into S. bovis did not affect formate and lactate production, which suggests that the activity of the pfl promoter is low. When the pfl promoter was replaced by the S. bovis ldh promoter, PFL was overexpressed, which caused an increase in the formate-to-lactate ratio. However, when PFL-AE was overexpressed, the formate-to-lactate ratio did not change, suggesting that PFL-AE was present at a level that was high enough to activate PFL. When both PFL-AE and PFL were overexpressed, the formate-to-lactate ratio further increased. It is conceivable that LDH activity is much higher than PFL activity, which may explain why the formate-to-lactate ratio is usually low.

* Corresponding author. Mailing address: Department of Life Science, College of Agriculture, Meiji University, Higashimita, Tama-ku, Kawasaki 214-8571, Japan. Phone: 81-44-934-7825. Fax: 81-44-934-7825. E-mail: hino{at}isc.meiji.ac.jp.

Applied and Environmental Microbiology, July 2002, p. 3352-3357, Vol. 68, No. 7
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.7.3352-3357.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Derzelle, S., Bolotin, A., Mistou, M.-Y., Rul, F. (2005). Proteome Analysis of Streptococcus thermophilus Grown in Milk Reveals Pyruvate Formate-Lyase as the Major Upregulated Protein. Appl. Environ. Microbiol. 71: 8597-8605 [Abstract] [Full Text]  
  • Asanuma, N., Yoshii, T., Hino, T. (2004). Molecular Characterization of CcpA and Involvement of This Protein in Transcriptional Regulation of Lactate Dehydrogenase and Pyruvate Formate-Lyase in the Ruminal Bacterium Streptococcus bovis. Appl. Environ. Microbiol. 70: 5244-5251 [Abstract] [Full Text]  


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